Regulatory T-cells (Treg) have been the focus of immunologic research due to their role in establishing tolerance for harmless antigens versus allowing immune responses against foes. Increased Treg frequencies measured by mRNA expression or protein synthesis of the Treg marker FOXP3 were found in various cancers, indicating that dysregulation of Treg levels contributes to tumor establishment. Furthermore, they constitute a key target of immunomodulatory therapies in cancer as well as transplantation settings. One core obstacle for understanding the role of Treg, thus far, is the inability of FOXP3 mRNA or protein detection methods to differentiate between Treg and activated T cells. These difficulties are aggravated by the technical demands of sample logistics and processing. Based on Treg-specific DNA demethylation within the FOXP3 locus, we present a novel method for monitoring Treg in human peripheral blood and solid tissues. We found that Treg numbers are significantly increased in the peripheral blood of patients with interleukin 2–treated melanoma and in formalin-fixed tissue from patients with lung and colon carcinomas. Conversely, we show that immunosuppressive therapy including therapeutic antibodies leads to a significant reduction of Treg from the peripheral blood of transplantation patients. In addition, Treg numbers are predictively elevated in the peripheral blood of patients with various solid tumors. Although our data generally correspond to data obtained with gene expression and protein-based methods, the results are less fluctuating and more specific to Treg. The assay presented here measures Treg robustly in blood and solid tissues regardless of conservation levels, promising fast screening of Treg in various clinical settings. [Cancer Res 2009;69(2):599–608]

The majority of transplants are derived from donors who suffered from brain injury. There is evidence that brain death causes inflammatory changes in the donor. To define the impact of brain death, we evaluated the gene expression of cytokines in human brain dead and ideal living donors and compared these data to organ function following transplantation. Hepatic tissues from brain dead (n = 32) and living donors (n = 26) were collected at the time of donor laparotomy. Additional biopsies were performed before organ preservation, at the time of transplantation and one hour after reperfusion. Cytokines were assessed by real-time reverse transcriptase–polymerase chain reaction (RT–PCR) and cytometric bead array. Additionally, immunohistological analysis of tissue specimens was performed. Inflammatory cytokines including IL-6, IL-10, TNF-α, TGF-β and MIP-1α were significantly higher in brain dead donors immediately after laparotomy compared to living donors. Cellular infiltrates significantly increased in parallel to the soluble cytokines IL-6 and IL-10. Enhanced immune activation in brain dead donors was reflected by a deteriorated I/R injury proven by elevated alanin-amino-transferase (ALT), aspartat-amino-transferase (AST) and bilirubin levels, increased rates of acute rejection and primary nonfunction. Based on our clinical data, we demonstrate that brain death and the events that precede it are associated with a significant upregulation of inflammatory cytokines and lead to a worse ischemia/reperfusion injury after transplantation.

Ischemia-reperfusion injury (IRI) constitutes a significant source of morbidity and mortality after orthotopic liver transplantation (OLT). The allograft is metabolically impaired during warm and cold ischemia and is further damaged by a paradox reperfusion injury after revascularization and reoxygenation. Short-term and long-term complications including post-reperfusion syndrome, delayed graft function, and immune activation have been associated with IRI. Due to the current critical organ shortage, extended criteria grafts are increasingly considered for transplantation, however, with an elevated risk to develop significant features of IRI. In recent years, ex vivo machine perfusion (MP) of the donor liver has witnessed significant advancements. Here, we describe the concept of hypothermic (oxygenated) machine perfusion (HMP/HOPE) approaches and highlight which allografts may benefit from this technology. This review also summarizes clinical applications and the main aspects of ongoing randomized controlled trials on hypothermic perfusion. The mechanistic aspects of IRI and hypothermic MP—which include tissue energy replenishment, optimization of mitochondrial function, and the reduction of oxidative and inflammatory damage following reperfusion—will be comprehensively discussed within the context of current preclinical and clinical evidence. Finally, we highlight novel trends and future perspectives in the field of hypothermic MP in the context of recent findings of basic and translational research.

A short period (1–2 h) of hypothermic oxygenated machine perfusion (HOPE) after static cold storage is safe and reduces ischemia‐reperfusion injury‐related complications after liver transplantation. Machine perfusion time is occasionally prolonged for logistical reasons, but it is unknown if prolonged HOPE is safe and compromises outcomes. We conducted a multicenter, observational cohort study of patients transplanted with a liver preserved by prolonged (≥4 h) HOPE. Postoperative biochemistry, complications, and survival were evaluated. The cohort included 93 recipients from 12 European transplant centers between 2014–2021. The most common reason to prolong HOPE was the lack of an available operating room to start the transplant procedure. Grafts underwent HOPE for a median (range) of 4:42 h (4:00–8:35 h) with a total preservation time of 10:50 h (5:50–20:50 h). Postoperative peak ALT was 675 IU/L (interquartile range 419–1378 IU/L). The incidence of postoperative complications was low, and 1‐year graft and patient survival were 94% and 88%, respectively. To conclude, good outcomes are achieved after transplantation of donor livers preserved with prolonged (median 4:42 h) HOPE, leading to a total preservation time of almost 21 h. These results suggest that simple, end‐ischemic HOPE may be utilized for safe extension of the preservation time to ease transplantation logistics.

Introduction: Colorectal carcinomas represent the third most common cause of cancer-related deaths in Germany. Although the incidence is significantly higher in men compared with women and gender is a well-established crucial factor for outcome in other diseases, detailed gender comparisons for colon cancer are lacking. Methods: This retrospective population-based cohort study included all patients diagnosed with colon cancer in Germany between 2000 and 2016 who were included in the common dataset of colorectal cancer patients from the quality conference of the German Cancer Society. We compared clinical, histopathological, and therapeutic characteristics as well as overall and recurrence-free survival. Results: A total of 185,967 patients were included in the study, of which 85,685 were female (46.1%) and 100,282 were male (53.9%). The proportion of women diagnosed with colon cancer decreased from 2000 to 2016 (f: 26.6 to 40.1%; m: 24.9 to 41.9%; p < 0.001), and the proportion of very old patients was especially high in women (f: 27.3%; m: 15.6%; p < 0.001). The localization in women was more right-sided (f: 45.0%, m: 36.7%; p < 0.001), and women had a higher tumor grading and a higher UICC stage (especially stage III nodal-positive) at diagnosis of primary colon cancer (UICC III: f: 22.7%, m: 21.0%; p < 0.001). We could detect a significantly better overall (hazard ratio: 0.853, lower 95%: 0.841, upper 95%: 0.864; p < 0.001) and recurrence-free survival (hazard ratio: 0.857, lower 95%: 0.845, upper 95%: 0.868; p < 0.001) in women compared with men, even though women received chemotherapy less frequently compared with men (f: 26.1%, m: 28.1%; p < 0.001). Conclusion: We could detect several variables that differed significantly between men and women regarding clinical, histopathological, therapeutic, and outcome factors. We believe that it is crucial to consider gender as a key factor in the diagnosis and treatment of colon cancer. Sex-specific diagnostic tools could lead to an earlier diagnosis of colon cancer in women, and ways to increase the rate of chemotherapy in women should be evaluated. Furthermore, we recommend stratifying randomized trials by gender.

To improve the quality of organs from brain-dead donors by assessing the influence of alternative strategies on the early behavior of kidneys after transplantation into unmodified hosts. Kidneys transplanted from living donors perform consistently better than those from cadaver sources. The authors have recently shown that donor brain death produces inflammatory changes in peripheral organs within hours, amplifies coincident ischemia–reperfusion injury, and accelerates acute and chronic rejection. Normalization of the graft by donor hormone treatment has hitherto been unsuccessful. A standardized rat model of brain death was used. Experimental groups included recipients of allogeneic grafts from living and brain-dead donors (F344→LEW). Donors were treated immediately after induction of brain death either with intravenous steroids, which block inflammatory cytokine release, or a soluble P-selectin glycoprotein ligand (sPSGL), which blocks initial selectin-mediated cellular adhesion. Kidney grafts were examined serially up to 10 days by morphology, immmunohistology, and reverse transcriptase–polymerase chain reaction. Overall survival of ummodified recipients of kidneys from brain-dead donors was significantly reduced versus living donors. Animals with organs from brain-dead donors that had received steroids or sPSGL survived significantly longer than those from untreated brain-dead donors. The intensity of ischemia–reperfusion injury and of acute rejection was reduced. Cellular infiltration and transcription of mRNA of representative proinflammatory mediators were diminished. Treatment of organ donors at the time of brain death markedly improves organ quality after kidney transplantation, upgrading it to that from a living donor.